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F4/80 Antibody Staining Protocol for Immunohistochemistry
Description: MCAP497 recognises the mouse F4/80 antigen, a 160kD glycoprotein expressed by murine macrophages. The antibody binds to macrophages from different sites including the peritoneal cavity, lung, spleen and thymus, to blood monocytes and to macrophages derived from bone marrow precursors in culture. The antigen is not expressed by lymphocytes or polymorphonuclear cells.
Primary Antibody
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Name: F4/80 Antibody |
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Clone: CI:A3-1, Rat anti-Mouse |
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Supplier: AbD Serotec |
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Catalog Number: MCAP497 |
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Dilution: 1:50 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed. |
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Incubation Time/Temp: 60 minutes/room temperature |
Antigen Retrieval
| Device: Incubator |
| Buffer/pH value: IHC-TekTM Proteinase K Solution (Cat# IW-1101) |
| Heat/Cool Temperature: 37 ºC/room temperature |
| Heat/Cool Time: 20 minutes/20 minutes |
Detection Methods
| Standard Method: ABC Method or LSAB Method |
| Enhanced Method: Polymeric Methods |
Chromogen Substrate
| Reagent: DAB |
| Incubation Time/Temperature: 1-3 minutes/room temperature |
Counterstain
| Reagent: Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Membrane/cytoplasmic |
| Images: Search image |
Additional Information:
| Tissue Type: Spleen, lung, thymus. |
| Fixation: Formalin fixed paraffin sections, or acetone fixed frozen sections |
| Positive Control: Spleen, lung, thymus. |
| Negative Control: Omit primary antibody, isotype control or absorption control |
| Blocking: 2-5% normal serum to reduce unspecific background staining; 0.5-3% H2O2 to block endogenous peroxidase activity; avidin/biotin to block endogenous biotin activity if necessary |
Notes:
No need to perform antigen retrieval on frozen sections |
References: